The mechanisms by which leptin regulates energy homeostasis are not fully understood. Human obesity is most often characterized by hyperleptinemia and by leptin resistance that yet has to be explained. A number of observations demonstrate that hypothalamic proopiomelanocortin (POMC) is an important mediator of leptin action in the brain. POMC-expressing neurons in the hypothalamus express the leptin receptor and hypothalamic POMC mRNA levels are positively regulated by leptin. The intracellular signaling mechanisms by which leptin regulate hypothalamic pomc gene expression are unknown and will be investigated in detail in this proposal. POMC is a precursor that encompasses a number of smaller peptides, including alpha-melanocyte-stimulating-hormone (alpha-MSH), which is generated via proteolytic processing by prohormone convertases (PC1 and PC2). Hypothalamic-derived alpha-MSH is a primary source of ligand for central melanocortin receptors and plays a crucial inhibitory role in central regulation of feeding and energy storage. We have new data demonstrating that leptin also stimulates expression of hypothalamic PC1 and PC2. This suggests that leptin can regulate hypothalamic POMC at both the transcriptional and posttranslational levels. We will investigate the coordinated regulation by leptin of hypothalamic POMC gene-transcription, biosynthesis and processing, and will characterize the post-translational modification and secretion of alpha-MSH. Since POMC neurons express melanocortin receptors and are targets of NPY/AgRP neurons, we will also investigate modulation of POMC expression and processing by these neuropeptides. In addition to transfection and in vivo studies, we will as a key model system utilize a unique and well-defined primary fetal culture of hypothalamic neurons. The physiological relevance of the melanocortin pathway in regulation of body weight is also testified by the mutations in this pathway causing obesity in humans and in rodents. The proposed studies will lead to a better understanding of the effects of leptin on hypothalamic POMC biology and are therefore likely to shed light on mechanisms of body-weight regulation and the development of obesity.